Structures and Functions of Membrane Antenna Proteins with High Resolution Solution and Solid State NMR

本研究的主要目的是建立高分辨率溶液核磁共振与固态核磁共振相结合的方法来阐明膜蛋白和色素配合物的结构和功能。我们选择了一种细菌膜天线和一种叶绿体天线作为研究对象。这种膜天线被称为LH1（光收集复合物1）或B870，其亚基（B820）由两个（α和β）多肽和活性位点的两个细菌叶绿素分子组成作为假基。我们已经建立了利用选择性同位素富集样品来阐明假体基团周围结构的方法。因此，我们用富集的同位素BCh1 a和正常亚基重构了LH1的B820亚基，或者用富集的同位素a和天然β亚基以及天然BCh1 a重构了LH1的B820亚基，并利用这些样品进行了溶液核磁共振分析。在这种情况下，富含^<13>C的色素细菌叶绿素和富含^2H的蛋白质可以聚焦细菌叶绿素和环境的信号。此外，我们还对这些样品广泛使用了多维核磁共振。这些研究揭示了在核磁共振方面阐明膜蛋白的方法。LH1的整个单元，即B870太大，无法用溶液核磁共振来研究，我们尝试充分利用固体高分辨率核磁共振来观察核磁共振。根据溶液核磁共振的知识对所得数据进行了分析。分配的信号告诉我们B870将由溶液状态下具有相似结构的亚基组成。多维溶液核磁共振和同位素富集为澄清氯仿/甲醇溶液中组分a和R多肽的结构提供了方法

Main object of the research is to establish the method to elucidate structures and functions of membrane proteins and pigment complexes in terms of high resolution solution nuclear magnetic resonance in conjunction with solid state NMR. For the target of the studies we have selected one of the bacterial membrane antenna, and chlorosome anntena. The membrane antenna called LH1 (light harvesting complex 1) or B870 has subunits (B820) composed of two (α and β) polypeptides and two bacteriochlorophyll molecultes in the active site as the prothetic groups. We have established the methods to elucidate the structures around the prothetic group with use of selectively isotope enriched samples. Thus, we reconstituted the subunit B820 of the LH1 with the isotope enrichied BCh1 a and the normal subumits, or the isotope enriched a and the natural β subunit and the natural BChl a and could resolve the structures with use these samples by the solution NMR. In this case the ^<13>C enriched pigment bacteriochlorophylls and the ^2H enriched proteins could focus the signal of the bacteriochlorophylls and the environment. Futhermore we have make extensive use of multidimensional NMR with these samples. These studies have disclosed the method to elucidate membrane proteins in terms of NMR. The whole unit of LH1, i.e., B870 is too large to be investigated by the solution NMR, we have tried to observe the NMR in full use of solid high resolution NMR. The obtained data have been analyzed in terms of the knowledge of the solution NMR. The assigned signal tells us the B870 would be composed of the subunits with similar structures in the solution states. The multi-dimensional solution NMR and the isotope enrichment have afforded the method to clarify the structure of the component a and R polypeptide in the chloroform/methanol solution